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Tumor metastasis remains the main cause of breast cancer-related deaths, especially delayed breast cancer distant metastasis. The current study assessed the frequency of CD44 - /CD24 -  breast cancer cells in 576 tissue specimens for associations with clinicopathological features and metastasis and investigated the underlying molecular mechanisms. The results indicated that higher frequency (≥19.5%) of CD44 - /CD24 -  cells was associated with delayed postoperative breast cancer metastasis. Furthermore, CD44 - /CD24 - triple negative breast cancer (TNBC) cells spontaneously converted into CD44 + /CD24 - cancer stem cells (CSCs) with properties similar to CD44 + /CD24 - CSCs from primary human breast cancer cells and parental TNBC cells in terms of stemness marker expression, self-renewal, differentiation, tumorigenicity, and lung metastasis in vitro and  in NOD/SCID mice . RNA sequencing identified several differentially expressed genes (DEGs) in newly converted CSCs and  RHBDL2 , one of the DEGs, expression was upregulated. More importantly,  RHBDL2  silencing inhibited the YAP1/USP31/NF-κB signaling and attenuated spontaneous CD44 - /CD24 -  cell conversion into CSCs and their mammosphere formation. These findings suggest that the frequency of CD44 - /CD24 -  tumor cells and  RHBDL2  may be valuable for prognosis of delayed breast cancer metastasis, particularly for TNBC.

Association of human breast cancer CD44 - /CD24 - cells with delayed distant metastasis